P309: IMAGE-BASED HIGH-CONTENT DRUG SENSITIVITY SCREENING IDENTIFIES CONVENTIONAL, EMERGING, AND POTENTIAL NOVEL THERAPEUTIC TARGETS IN HIGH-RISK ADULT ACUTE LYMPHOBLASTIC LEUKEMIA
Authors: H. Bell, M. Singh, H. Blair, A. Poll, E. Law, O. Heidenreich, F. van Delft, A. Moorman, J. Lunec, J. Irving
Journal: HemaSphere (2022)

Background: Despite concerted efforts to optimize and intensify treatment approaches in adult acute lymphoblastic leukemia (ALL) more than half of patients still succumb to their disease within 5 years of diagnosis, emphasizing the limits of current therapy and the critical requirement for more effective and safer therapeutics. Patient-derived xenograft (PDX) ALL models represent reliably predictive preclinical models and allow in vivo amplification of patient material in quantities sufficient for high-throughput screening applications. Aims: To identify novel and effective therapies for high-risk ALL using an in vitro co-culture model of PDX ALL blasts and mesenchymal stem cells (MSCs) by high-content screening of >1200 pharmacologically-active small-molecule drug compounds in various stages of clinical development. Methods: PDX leukemic cells derived from seven patients with high-risk ALL, including relapsed Philadelphia-positive B-ALL, low hypodiploid B-ALL, and ABL-class T-ALL, were used for the drug screen. ALL cells were co-cultured with hTERT-immortalized MSCs to support survival of the ALL blasts during a 96 hour short-term culture with the drug library compounds (final concentration, 1mM) in 384-well format. Drug sensitivity was assessed using an automated, high-throughput, fluorescence image-based microscopy pipeline which identifies and discriminates live ALL cells and MSCs using random forest machine learning algorithms based on cellular nuclear staining. Results: Drug sensitivity profiles revealed a wide diversity of responses to the library compounds reflected by different “hit” rates (4.5-7.4%), where “hit” is defined as compounds reducing cell survival by more than 50%. Despite this heterogeneity in response, 59 unique compounds were identified with broad anti-leukemic activity; that is, “hits” in 4/7 PDX samples. Concurrent image analysis of MSCs in the co-culture system revealed most hit compounds did not affect MSC numbers relative to vehicle-treated controls (n=50/59), indicating a possible therapeutic window for targeting leukemic cells specifically. Several hit compounds (n=14) were drugs or from drug classes which form part of conventional ALL treatment regimens — including anthracyclines, glucocorticoids, and nucleoside analogues — validating the screening approach for successful identification of anti-leukemic compounds. Drug classes with recently recognized and emerging preclinical anti-leukemic activity were also identified including cyclin-dependent kinase (CDK) inhibitors (n=5), cell cycle-related kinase inhibitors (n=6), bromodomain and extra-terminal domain (BET) inhibitors (n=4), histone deacetylase family inhibitors (n=3), and nicotinamide phosphoribosyltransferase (NAMPT) inhibitors (n=3). Nine hit compounds elicited significant reductions in cell survival consistently across the panel of PDX samples (7/7), providing some evidence towards anti-leukemic activity independent of cytogenetic background. These included pan-CDK family inhibitors alvocidib, CGP60474, and roscovitine-derived CR8. Additionally, the selective CDK (-1, -2, -9) inhibitor AZD 5438 exhibited anti-leukemic activity in all samples (n=6/7), bar the Philadelphia-positive sample. Summary/Conclusion: We provide evidence that high-content drug screening in a co-culture system is an effective strategy to identify clinically-relevant novel and emerging therapeutic targets in high-risk ALL. Our analyses promote a strong rationale for exploring the targeting of CDKs as a potential pan-effective approach in high-risk adult ALL.